Physiological and genomic insights into a psychrotrophic drought-tolerant bacterial consortium for crop improvement in cold, semiarid regions.

The agricultural land in the Indian Himalayan region (IHR) is susceptible to various spells of snowfall, which can cause nutrient leaching, low temperatures, and drought conditions. The current study, therefore, sought an indigenous psychrotrophic plant growth-promoting (PGP) bacterial inoculant with the potential to alleviate crop productivity under cold and drought stress. Psychrotrophic bacteria preisolated from the night-soil compost of the Lahaul Valley of northwestern Himalaya were screened for phosphate (P) and potash (K) solubilization, nitrogen fixation, indole acetic acid (IAA) production, siderophore and HCN production) in addition to their tolerance to drought conditions for consortia development. Furthermore, the effects of the selected consortium on the growth and development of wheat (Triticum aestivum L.) and maize (Zea mays L.) were assessed in pot experiments under cold semiarid conditions (50 % field capacity). Among 57 bacteria with P and K solubilization, nitrogen fixation, IAA production, siderophore and HCN production, Pseudomonas protegens LPH60, Pseudomonas atacamensis LSH24, Psychrobacter faecalis LUR13, Serratia proteamaculans LUR44, Pseudomonas mucidolens LUR70, and Glutamicibacter bergerei LUR77 exhibited tolerance to drought stress (-0.73 MPa). The colonization of wheat and maize seeds with these drought-tolerant PGP strains resulted in a germination index >150, indicating no phytotoxicity under drought stress. Remarkably, a particular strain, Pseudomonas sp. LPH60 demonstrated antagonistic activity against three phytopathogens Ustilago maydis, Fusarium oxysporum, and Fusarium graminearum. Treatment with the consortium significantly increased the foliage (100 % and 160 %) and root (200 % and 133 %) biomasses of the wheat and maize plants, respectively. Furthermore, whole-genome sequence comparisons of LPH60 and LUR13 with closely related strains revealed genes associated with plant nutrient uptake, phytohormone synthesis, siderophore production, hydrogen cyanide (HCN) synthesis, volatile organic compound production, trehalose and glycine betaine transport, cold shock response, superoxide dismutase activity, and gene clusters for nonribosomal peptide synthases and polyketide synthetases. With their PGP qualities, biocontrol activity, and ability to withstand environmental challenges, the developed consortium represents a promising cold- and drought-active PGP bioinoculant for cereal crops grown in cold semiarid regions.

RedOx regulation of LRRK2 kinase activity by active site cysteines.

Mutations of the human leucine-rich repeat kinase 2 (LRRK2) have been associated with both, idiopathic and familial Parkinson's disease (PD). Most of these pathogenic mutations are located in the kinase domain (KD) or GTPase domain of LRRK2. In this study we describe a mechanism in which protein kinase activity can be modulated by reversible oxidation or reduction, involving a unique pair of adjacent cysteines, the "CC" motif. Among all human protein kinases, only LRRK2 contains this "CC" motif (C2024 and C2025) in the Activation Segment (AS) of the kinase domain. In an approach combining site-directed mutagenesis, biochemical analyses, cell-based assays, and Gaussian accelerated Molecular Dynamics (GaMD) simulations we could attribute a role for each of those cysteines. We employed reducing and oxidizing agents with potential clinical relevance to investigate effects on kinase activity and microtubule docking. We find that each cysteine gives a distinct contribution: the first cysteine, C2024, is essential for LRRK2 protein kinase activity, while the adjacent cysteine, C2025, contributes significantly to redox sensitivity. Implementing thiolates (R-S-) in GaMD simulations allowed us to analyse how each of the cysteines in the "CC" motif interacts with its surrounding residues depending on its oxidation state. From our studies we conclude that oxidizing agents can downregulate kinase activity of hyperactive LRRK2 PD mutations and may provide promising tools for therapeutic strategies.

Acute demyelinating encephalomyelitis in a child with pulmonary tuberculosis.

Tuberculosis is a leading cause of mortality in children worldwide. One of the greatest challenges in its management is the difficulty of diagnosis as the manifestations are non-specific and often mimic other illnesses. Neurological infection occurs in approximately 1% of patients diagnosed with tuberculosis, and usually takes the form of tuberculous meningitis or tuberculoma. An 11-year-old girl who was diagnosed with acute disseminated encephalomyelitis, a rare immunological manifestation of tuberculosis, is presented. She recovered completely after a course of high-dose systemic corticosteroids in addition to anti-tuberculosis treatment. Considering the immense burden of this infectious disease, recognition and understanding of the uncommon manifestations are important to enable appropriate and timely treatment.Abbreviations: ADEM: acute disseminated encephalomyelitis; ATT: anti-tuberculosis therapy; CBNAAT: cartridge-based nucleic acid amplification test; CNS: central nervous system; CSF: cerebrospinal fluid; CT: computed tomography; FLAIR: fluid attenuated inversion recovery; IFN: interferon; MRI: magnetic resonance imaging; MTB: Mycobacterium tuberculosis; TB: tuberculosis; TNF: tumour necrosis factor.

Addressing lanthanum toxicity in plants: Sources, uptake, accumulation, and mitigation strategies.

Lanthanum (La), the second most abundant rare earth element (REE) is emerging as an environmental issue, with the potential to impact ecosystems and human health. Major sources of soil contamination by La include agricultural, and industrial activities. Lanthanum is non-essential for plant growth but accumulates in various plant parts. The uptake of La by plants is intricately influenced by various factors such as soil pH, redox potential, cation exchange capacity, presence of organic acids and rhizosphere composition. These factors significantly impact the availability and absorption of La ions. Lanthanum impact on plants depends on soil characteristics, cultivated species, developmental stage, La concentration, treatment period, and growth conditions. Excessive La concentrations affect cell division, DNA structure, nutrient uptake, and photosynthesis and induce toxicity symptoms. Plants employ detoxification mechanisms like vacuolar sequestration, osmolyte synthesis, and antioxidant defense system. However, higher concentrations of La can overwhelm these defense mechanisms, leading to adverse effects on plant growth and development. Further, accumulation of La in plants increases the risk for human exposure. Strategies to mitigate La toxicity are, therefore, vital for ecosystem protection. The application of phytoremediation, supplementation, chelation, amendments, and biosorption techniques contributes to the mitigation of La toxicity. This review provides insights into La sources, uptake, toxicity, and alleviation strategies in plants. Identifying research gaps and discussing advancements aims to foster a holistic understanding and develop effective strategies for protecting plant health and ecosystem resilience against La contamination.

The efficient applications of native flora for phytorestoration of mine tailings: a pan-global survey.

Mine tailings are the discarded materials resulting from mining processes after minerals have been extracted. They consist of leftover mineral fragments, excavated land masses, and disrupted ecosystems. The uncontrolled handling or discharge of tailings from abandoned mine lands (AMLs) poses a threat to the surrounding environment. Numerous untreated mine tailings have been abandoned globally, necessitating immediate reclamation and restoration efforts. The limited feasibility of conventional reclamation methods, such as cost and acceptability, presents challenges in reclaiming tailings around AMLs. This study focuses on phytorestoration as a sustainable method for treating mine tailings. Phytorestoration utilizes existing native plants on the mine sites while applying advanced principles of environmental biotechnology. These approaches can remediate toxic elements and simultaneously improve soil quality. The current study provides a global overview of phytorestoration methods, emphasizing the specifics of mine tailings and the research on native plant species to enhance restoration ecosystem services.

Nanowonders in agriculture: Unveiling the potential of nanoparticles to boost crop resilience to salinity stress.

Soil salinization significantly affects crop production by reducing crop quality and decreasing yields. Climate change can intensify salinity-related challenges, making the task of achieving global food security more complex. To address the problem of elevated salinity stress in crops, nanoparticles (NPs) have emerged as a promising solution. NPs, characterized by their small size and extensive surface area, exhibit remarkable functionality and reactivity. Various types of NPs, including metal and metal oxide NPs, carbon-based NPs, polymer-based NPs, and modified NPs, have displayed potential for mitigating salinity stress in plants. However, the effectiveness of NPs application in alleviating plant stress is dependent upon multiple factors, such as NPs size, exposure duration, plant species, particle composition, and prevailing environmental conditions. Moreover, alterations to NPs surfaces through functionalization and coating also play a role in influencing plant tolerance to salinity stress. NPs can influence cellular processes by impacting signal transduction and gene expression. They counteract reactive oxygen species (ROS), regulate the water balance, enhance photosynthesis and nutrient uptake and promote plant growth and yield. The objective of this review is to discuss the positive impacts of diverse NPs on alleviating salinity stress within plants. The intricate mechanisms through which NPs accomplish this mitigation are also discussed. Furthermore, this review addresses existing research gaps, recent breakthroughs, and prospective avenues for utilizing NPs to combat salinity stress.

Role of the leucine-rich repeat protein kinase 2 C-terminal tail in domain cross-talk.

Leucine-rich repeat protein kinase 2 (LRRK2) is a multi-domain protein encompassing two of biology's most critical molecular switches, a kinase and a GTPase, and mutations in LRRK2 are key players in the pathogenesis of Parkinson's disease (PD). The availability of multiple structures (full-length and truncated) has opened doors to explore intra-domain cross-talk in LRRK2. A helix extending from the WD40 domain and stably docking onto the kinase domain is common in all available structures. This C-terminal (Ct) helix is a hub of phosphorylation and organelle-localization motifs and thus serves as a multi-functional protein : protein interaction module. To examine its intra-domain interactions, we have recombinantly expressed a stable Ct motif (residues 2480-2527) and used peptide arrays to identify specific binding sites. We have identified a potential interaction site between the Ct helix and a loop in the CORB domain (CORB loop) using a combination of Gaussian accelerated molecular dynamics simulations and peptide arrays. This Ct-Motif contains two auto-phosphorylation sites (T2483 and T2524), and T2524 is a 14-3-3 binding site. The Ct helix, CORB loop, and the CORB-kinase linker together form a part of a dynamic 'CAP' that regulates the N-lobe of the kinase domain. We hypothesize that in inactive, full-length LRRK2, the Ct-helix will also mediate interactions with the N-terminal armadillo, ankyrin, and LRR domains (NTDs) and that binding of Rab substrates, PD mutations, or kinase inhibitors will unleash the NTDs.

Mitigating cadmium accumulation and toxicity in plants: The promising role of nanoparticles.

Cadmium (Cd) is a highly toxic heavy metal that adversely affects humans, animals, and plants, even at low concentrations. It is widely distributed and has both natural and anthropogenic sources. Plants readily absorb and distribute Cd in different parts. It may subsequently enter the food chain posing a risk to human health as it is known to be carcinogenic. Cd has a long half-life, resulting in its persistence in plants and animals. Cd toxicity disrupts crucial physiological and biochemical processes in plants, including reactive oxygen species (ROS) homeostasis, enzyme activities, photosynthesis, and nutrient uptake, leading to stunted growth and reduced biomass. Although plants have developed defense mechanisms to mitigate these damages, they are often inadequate to combat high Cd concentrations, resulting in yield losses. Nanoparticles (NPs), typically smaller than 100 nm, possess unique properties such as a large surface area and small size, making them highly reactive compared to their larger counterparts. NPs from diverse sources have shown potential for various agricultural applications, including their use as fertilizers, pesticides, and stress alleviators. Recently, NPs have emerged as a promising strategy to mitigate heavy metal stress, including Cd toxicity. They offer advantages, such as efficient absorption by crop plants, the reduction of Cd uptake, and the enhancement of mineral nutrition, antioxidant defenses, photosynthetic parameters, anatomical structure, and agronomic traits in Cd-stressed plants. The complex interaction of NPs with calcium ions (Ca2+), intracellular ROS, nitric oxide (NO), and phytohormones likely plays a significant role in alleviating Cd stress. This review aims to explore the positive impacts of diverse NPs in reducing Cd accumulation and toxicity while investigating their underlying mechanisms of action. Additionally, it discusses research gaps, recent advancements, and future prospects of utilizing NPs to alleviate Cd-induced stress, ultimately promoting improved plant growth and yield.

Laser Light as an Emerging Method for Sustainable Food Processing, Packaging, and Testing.

In this review article, we systematically investigated the diverse applications of laser technology within the sphere of food processing, encompassing techniques such as laser ablation, microbial inactivation, state-of-the-art food packaging, and non-destructive testing. With a detailed exploration, we assess the utility of laser ablation for the removal of surface contaminants from foodstuffs, while also noting the potential financial and safety implications of its implementation on an industrial scale. Microbial inactivation by laser shows promise for reducing the microbial load on food surfaces, although concerns have been raised about potential damage to the physio-characteristics of some fruits. Laser-based packaging techniques, such as laser perforation and laser transmission welding, offer eco-friendly alternatives to traditional packaging methods and can extend the shelf life of perishable goods. Despite the limitations, laser technology shows great promise in the food industry and has the potential to revolutionize food processing, packaging, and testing. Future research needs to focus on optimizing laser equipment, addressing limitations, and developing mathematical models to enhance the technology's uses.

Picolinic acid is a broad-spectrum inhibitor of enveloped virus entry that restricts SARS-CoV-2 and influenza A virus in vivo.

The COVID-19 pandemic highlights an urgent need for effective antivirals. Targeting host processes co-opted by viruses is an attractive antiviral strategy with a high resistance barrier. Picolinic acid (PA) is a tryptophan metabolite endogenously produced in mammals. Here, we report the broad-spectrum antiviral activity of PA against enveloped viruses, including severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), influenza A virus (IAV), flaviviruses, herpes simplex virus, and parainfluenza virus. Mechanistic studies reveal that PA inhibits enveloped virus entry by compromising viral membrane integrity, inhibiting virus-cellular membrane fusion, and interfering with cellular endocytosis. More importantly, in pre-clinical animal models, PA exhibits promising antiviral efficacy against SARS-CoV-2 and IAV. Overall, our data establish PA as a broad-spectrum antiviral with promising pre-clinical efficacy against pandemic viruses SARS-CoV-2 and IAV.

Alleviation of chromium stress in plants using metal and metal oxide nanoparticles.

Chromium (Cr), one of the hazardous pollutants, exists predominantly as Cr(VI) and Cr(III) in the environment. Cr(VI) is more toxic than Cr(III) due to its high mobility and solubility. Elevated levels of Cr in agricultural soil due to various anthropogenic activities cause Cr accumulation in plants, resulting in a significant reduction in plant yield and quality due to Cr-induced physiological, biochemical and molecular alterations. It can infiltrate the food chain through crop plants and cause harmful effects in humans via biomagnification. Cr(VI) is linked to cancer in humans. Therefore, mitigation strategies are required to remediate Cr-polluted soils and limit its accumulation in plants for safe food production. Recent research on metal and metal oxide nanoparticles (NPs) has shown that they can effectively reduce Cr accumulation and phytotoxicity. The effects of these NPs are influenced by their type and dose, exposure method, plant species and experimental settings. In this review, we present an up-to-date compilation and comprehensive analysis of the existing literature regarding the process of uptake and distribution of Cr and impact and potential mechanisms of metal and metal oxide nanoparticles led mitigation of Cr-induced stress in plants. We have also discussed recent developments, existing research gaps and future research directions in the field of Cr stress mitigation by NPs in plants. Overall, this review can provide valuable insights in reducing Cr accumulation and toxicity using metal and metal oxide nanoparticles, thereby promoting safe and sustainable cultivation of food and phytostabilization of Cr-polluted soil.

Lens Coloboma: A Rare Association of Congenital Rubella Syndrome.

Congenital rubella syndrome (CRS) may affect all ocular structures in general, either in isolation or in combination. Typical ocular complications in CRS include cataracts, microcornea, microphthalmia, glaucoma, nystagmus, and retinopathy. We report a case of a four-year-old girl who presented with bilateral total cataracts with sensory nystagmus, a poorly dilating pupil, iris hypoplasia, high axial myopia in the right eye (RE), and a retinal detachment in the left eye (LE). The systemic evaluation revealed microcephaly with an associated patent ductus arteriosus (PDA) and mild pulmonary arterial hypertension (PAH). Based on these findings, the child was diagnosed with clinically confirmed CRS. The child was taken up for right-eye cataract surgery. Intra-operatively, a lens coloboma involving the temporal equator of the lens along with a highly tessellated fundus was noted. Two weeks post-cataract surgery, the child had a best corrected visual acuity (BCVA) of 6/24. However, four weeks after the surgery, the child developed total rhegmatogenous retinal detachment in the right eye, for which pars plana vitrectomy with endolaser and silicon oil tamponade was done. Four weeks later, the child's BCVA in the right eye was 6/36, which was maintained until the last follow-up of four months. Lens coloboma may be isolated or may occur in association with chorioretinal coloboma. Ours is the first case of unilateral atypical lens coloboma associated with high myopia and bilateral cataracts in a patient with congenital rubella syndrome. Lens colobomas with high myopia have not been reported previously as ocular associations of CRS. Our case highlights that in children with CRS presenting with unilateral or bilateral congenital cataracts, the possibility of lenticular coloboma as a coexistent association should be kept in mind while taking these cases for cataract surgery.

Challenges in the Classification of Oral Submucous Fibrosis and Proposing a New Classification Based on Systematic Review of Literature.

Aims and Objectives: Oral submucous fibrosis is one of the common oral potentially malignant disorders that can result in severe morbidity. Due to its widespread involvement in the oral cavity and high risk of malignant transformation, the diagnosis and treatment of this disease at an early stage is essential to prevent further complications. This research was carried out to review various classification systems of oral submucous fibrosis documented in the literature to date, including their benefits and drawbacks, thus focusing on a need to discover some reliable classification systems. Materials and Methods: An electronic search of the published English literature was performed without publication year limitation in PubMed/Medline, Science Direct, Web of Science, Google Scholar, and Scopus databases using keywords like ('Oral submucous fibrosis' OR 'Oral submucous fibroses'), AND ('Classification' OR 'Grade' OR 'Stage'), AND ('Clinical', OR/ AND 'Histological', OR/AND 'Functional') following PRISMA guidelines. A manual search of all related Dental and Medical journals was also performed. We also checked the reference lists of the relevant articles for any other possible information on the subject. Results: The search strategy revealed 31 relevant articles and it could be appreciated that oral submucous fibrosis has been classified in seven different ways. Each system is enclosed with its own limitations and benefits. Conclusion: From this research, it can be concluded that despite the existence of several classification systems for oral submucous fibrosis, at present, no classification scheme is considered to be reliable that could help in the accurate assessment of the progression of the disease and classifying oral submucous fibrosis still remains a challenging task for clinicians, surgeons, and pathologists. We have postulated a proposed new classification system based on our literature research, but still robust research is needed in this aspect.

Capturing the domain crosstalk in full length LRRK2 and LRRK2RCKW.

LRRK2 is a multi-domain protein with three catalytically inert N-terminal domains (NtDs) and four C-terminal domains, including a kinase and a GTPase domain. LRRK2 mutations are linked to Parkinson's Disease. Recent structures of LRRK2RCKW and a full-length inactive LRRK2 (fl-LRRK2INACT) monomer revealed that the kinase domain drives LRRK2 activation. The LRR domain and also an ordered LRR- COR linker, wrap around the C-lobe of the kinase domain and sterically block the substrate binding surface in fl-LRRK2INACT. Here we focus on the crosstalk between domains. Our biochemical studies of GTPase and kinase activities of fl-LRRK2 and LRRK2RCKW reveal how mutations influence this crosstalk differently depending on the domain borders investigated. Furthermore, we demonstrate that removing the NtDs leads to altered intramolecular regulation. To further investigate the crosstalk, we used Hydrogen-Deuterium exchange Mass Spectrometry (HDX-MS) to characterize the conformation of LRRK2RCKW   and Gaussian Accelerated Molecular Dynamics (GaMD) to create dynamic portraits of fl-LRRK2 and LRRK2RCKW. These models allowed us to investigate the dynamic changes in wild type and mutant LRRK2s. Our data show that the a3ROC helix, the Switch II motif in the ROC domain, and the LRR-ROC linker play crucial roles in mediating local and global conformational changes. We demonstrate how these regions are affected by other domains in fl-LRRK2 and LRRK2RCKW and show how unleashing of the NtDs as well as PD mutations lead to changes in conformation and dynamics of the ROC and kinase domains which ultimately impact kinase and GTPase activities. These allosteric sites are potential therapeutic targets.

Capturing Differences in the Regulation of LRRK2 Dynamics and Conformational States by Small Molecule Kinase Inhibitors.

Mutations in the human leucine rich repeat protein kinase-2 (LRRK2) create risk factors for Parkinson's disease, and pathological functions of LRRK2 are often correlated with aberrant kinase activity. Past research has focused on developing selective LRRK2 kinase inhibitors. In this study, we combined enhanced sampling simulations with HDX-MS to characterize the inhibitor-induced dynamic changes and the allosteric communications within the C-terminal domains of LRRK2, LRRK2RCKW. We find that the binding of MLi-2 (a type I kinase inhibitor) stabilizes a closed kinase conformation and reduces the global dynamics of LRRK2RCKW, leading to a more compact LRRK2RCKW structure. In contrast, the binding of Rebastinib (a type II kinase inhibitor) stabilizes an open kinase conformation, which promotes a more extended LRRK2RCKW structure. By probing the distinct effects of the type I and type II inhibitors, key interdomain interactions are found to regulate the communication between the kinase domain and the GTPase domain. The intermediate states revealed in our simulations facilitate the efforts toward in silico design of allosteric modulators that control LRRK2 conformations and potentially mediate the oligomeric states of LRRK2 and its interactions with other proteins.

Preclinical evaluation of ADVM-062, a novel intravitreal gene therapy vector for the treatment of blue cone monochromacy.

Blue cone monochromacy (BCM) is a rare X-linked retinal disease characterized by the absence of L- and M-opsin in cone photoreceptors, considered a potential gene therapy candidate. However, most experimental ocular gene therapies utilize subretinal vector injection which would pose a risk to the fragile central retinal structure of BCM patients. Here we describe the use of ADVM-062, a vector optimized for cone-specific expression of human L-opsin and administered using a single intravitreal (IVT) injection. Pharmacological activity of ADVM-062 was established in gerbils, whose cone-rich retina naturally lacks L-opsin. A single IVT administration dose of ADVM-062 effectively transduced gerbil cone photoreceptors and produced a de novo response to long-wavelength stimuli. To identify potential first-in-human doses we evaluated ADVM-062 in non-human primates. Cone-specific expression of ADVM-062 in primates was confirmed using ADVM-062.myc, a vector engineered with the same regulatory elements as ADVM-062. Enumeration of human OPN1LW.myc-positive cones demonstrated that doses ≥3 × 1010 vg/eye resulted in transduction of 18%-85% of foveal cones. A Good Laboratory Practice (GLP) toxicology study established that IVT administration of ADVM-062 was well tolerated at doses that could potentially achieve clinically meaningful effect, thus supporting the potential of ADVM-062 as a one-time IVT gene therapy for BCM.

Sub-gingival delivery of simvastatin and rosuvastatin for treatment of chronic periodontitis with diabetes mellitus: A randomized controlled clinical-radiographic pilot study.

Background: Statins are lipid-lowering medications that work by blocking rate-limiting enzyme in cholesterol formation. In patients with Chronic periodontitis (CP) and Diabetes mellitus (DM), subgingival delivery of simvastatin (SMV) and rosuvastatin (RSV) have demonstrated to have bone-stimulatory and anti-inflammatory properties. The current study intended to assess and compare the efficacy of sub-gingivally delivered SMV gel and RSV gel as an adjunctive medication to scaling and root planing (SRP) in the management of intrabony defects in CP patients with type 2 DM. Methods: 30 patients with CP and type 2 DM were classified into three treatment groups - SRP + placebo, SRP +1.2% SMV and SRP +1.2% RSV. Clinical parameters: site-specific plaque index, modified sulcus bleeding index (mSBI), pocket probing depth (PPD), and relative attachment level (RAL) were documented at baseline, 3 and 6 months and radiographic parameter: intrabony defect depth (IBD) at baseline and 6 months post-treatment. Results: - LDD of 1.2% SMV and 1.2% RSV demonstrated greater clinical and radiographic improvement than placebo, the improvement being statistically significant for PI, mSBI, and PPD for 1.2% SMV and statistically significant for all clinical and radiological parameters for the 1.2% RSV. 1.2% RSV demonstrated greater IBD fill and RAL gain than 1.2% SMV. Conclusion: - Localized sub-gingival delivery of statins was beneficial in the treatment intrabony defects in patients with CP and well-controlled type 2 DM. IBD fill and RAL gain were higher with 1.2% RSV than with 1.2% SMV.

Novel LRR-ROC Motif That Links the N- and C-terminal Domains in LRRK2 Undergoes an Order-Disorder Transition Upon Activation.

Mutations in LRRK2, a large multi-domain protein kinase, create risk factors for Parkinson's Disease (PD). LRRK2 has seven well-folded domains that include three N-terminal scaffold domains (NtDs) and four C-terminal domains (CtDs). In full-length inactive LRRK2 there is an additional well-folded motif, the LRR-ROC Linker, that lies between the NtDs and the CtDs. This motif, which is stabilized by hydrophobic residues in the LRR and ROC/COR-A domains, is anchored to the C-Lobe of the kinase domain. The LRR-ROC Linker becomes disordered when the NtDs are unleashed from the CtDs following activation by Rab29 or by various PD mutations. A key residue within the LRR-ROC Linker, W1295, sterically blocks access of substrate proteins. The W1295A mutant blocks cis-autophosphorylation of S1292 and reduces phosphorylation of heterologous Rab substrates. GaMD simulations show that the LRR-Linker motif, P + 1 loop and the inhibitory helix in the DYGψ motif are very stable. Finally, in full-length inactive LRRK2 ATP is bound to the kinase domain and GDP:Mg to the GTPase/ROC domain. The fundamentally different mechanisms for binding nucleotide (G-Loop vs P-Loop) are captured by these GaMD simulations. In this model, where ATP binds with low affinity (µM range) to N-Lobe capping residues, the known auto-phosphorylation sites are located in the space that is sampled by the flexible phosphates thus providing a potential mechanism for cis-autophosphorylation.