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Biomedical subjects

K Prasad

Publications and source records attributed to K Prasad.

At least 73 records · Page 4Linked to original sources
PubMed1999

Protection of coronary angioplasty-induced oxidative stress by Isovue used during angioplasty.

BACKGROUND: Oxygen free radicals (OFRs) have been implicated in ischemic-reperfusion cardiac injury. Use of percutaneous transluminal coronary angioplasty (PTCA) has created renewed interest in salvation of ischemic myocardium. The PTCA procedure is similar to the ischemia-reperfusion model. It is possible that OFRs are increased following PTCA. However, OFR-related cardiac complications are uncommon and the evidence for lipid peroxidation is conflicting. PATIENTS AND METHODS: In this study the levels of plasma malondialdehyde, OFR-producing activity of polymorphonuclear leukocytes (PMNL-CL) and blood antioxidant enzymes (superoxide dismutase, catalase and glutathione peroxidase) were measured in peripheral venous blood of 50 consecutive patients with stable angina undergoing elective PTCA. The ability of Isovue (used during PTCA) and of streptokinase (used during thrombolysis) to scavenge OH in the high performance liquid chromatography method and to reduce OH-induced lipid peroxidation were also assessed. Patients were divided into three groups: group 1, single vessel PTCA; group 2, two or more vessel PTCA; and group 3, combined single and multivessel PTCA. RESULTS: The results indicated that there was an increase in PMNL-CL (22% to 44%) and a decrease in plasma malondialdehyde (33% to 40%) at 60 mins following PTCA. The activity of antioxidant enzymes remained unaltered. Isovue scavenged OH in a concentration-dependent manner and was complete at a concentration below that used in patients. Streptokinase, on the other hand, was ineffective in scavenging OH. CONCLUSIONS: These results suggest that, in spite of increased production of OFR by polymorphonuclear leukocytes and unaltered activity of antioxidant enzymes, lipid peroxidation decreased. Lack of lipid peroxidation may have been due to the OH-scavenging property of Isovue. The observed differences in OFR-related complications between PTCA and thrombolytic therapy may have been due to the antioxidant activity of Isovue.

Adult↗
PubMed1998

QT dispersion and risk factors for sudden cardiac death in patients with hypertrophic cardiomyopathy.

This study examines the relation of QT dispersion (QTd) on a surface electrocardiogram (ECG) to clinical features and established risk factors of sudden cardiac death (SCD) in patients with hypertrophic cardiomyopathy (HC). One hundred fifty-six consecutive patients with HC (91 men, mean age 41+/-15 years, range 7 to 79) and 72 normal subjects (41 men, mean age 39+/-9 years, range 20 to 60) were studied. Standard 12-lead ECGs were recorded from each subject using a MAC VU electrocardiograph. Patients with nonsinus rhythm, atrioventricular conduction block, QRS duration > 120 ms, age < 15 years, and low amplitude T waves were excluded from the analysis (n=51). Another 22 patients who were receiving amiodarone and/or sotalol therapy were also excluded. QT interval and QTd were measured using automated analysis in the remaining 83 patients (46 men, age 40+/-14 years, range 16 to 76). QT interval (406+/-38 ms), QTc interval (432+/-27 ms), and QTd (43+/-25 ms) were significantly greater in patients with HC than in normal controls (386+/-31 ms, 404+/-16 ms, 26+/-16 ms, respectively) (p <0.0001). QTd was significantly greater in patients with HC with chest pain compared with asymptomatic or mildly symptomatic patients (50+/-28 ms vs 37+/-20 ms, p=0.02). Increased QTd was found in patients with dyspnea New York Heart Association functional classes II/III than in those with dyspnea New York Heart Association functional class I (50+/-27 ms vs 38+/-22 ms, p=0.04). QTd was weakly correlated with maximum left ventricular wall thickness (r=0.228, p=0.038). No significant association was found between QTd and any risk factors for SCD. Thus, patients with HC have increased QTd. The QTd correlates with symptomatic status. Assessment of QTd might provide complementary clinical characterization of patients with HC but its relation to SCD remains uncertain.

Adolescent↗
PubMed1998

Probing the phosphoinositide binding site of the clathrin assembly protein AP-2 with photoaffinity labels.

The relative binding specificities of the subunitsof bovine assembly protein AP-2 for the phosphatidylinositol polyphosphates (PtdInsPn) and inositol polyphosphates (InsPn) were determined by photoaffinitylabeling. Three types of benzophenone-containing photoprobes were employed: (i) the water-solubleP-1- or P-2-tethered p-benzoyldihydrocinnamoyl-InsPn (BZDC-InsPn) analogs, (ii) P-1-linked phosphotriester PtdInsPn analogs that sampled the interface between the water and lipid phases, and (iii) sn-1-O-acyl-linked PtdInsPn analogs that interacted with proteins penetrating the bilayer. The InsPn and PtdInsPn probes bind with highest selectivity and affinity to the two alpha subunit isoforms, with certain probes and conditions resulting in strong labeling of the 50-kDa mu subunit. Three main conclusions were reached: (i) head group recognition predominated over acyl chain recognition, (ii) the PtdInsPn binding site of alpha-AP-2 prefers more highly phosphorylated species, and (iii) the protein-acyl chain interactions showed high capacity but low selectivity.

Adaptor Proteins, Vesicular Transport↗
PubMed1998

Fumarate modulates bacterial flagellar rotation by lowering the free energy difference between the clockwise and counterclockwise states of the motor.

Switching flagellar rotation from one direction to another is an essential part of bacterial chemotaxis. Fumarate has been shown to possess the capacity to restore to flagella of cytoplasm-free, CheY-containing bacterial envelopes the ability to switch directions and to increase the probability of reversal in intact cells. Neither the target of fumarate action nor the mechanism of function is known. To distinguish between the two potential targets of fumarate, the response regulator CheY and the flagellar switch-motor complex, we compared flagellar rotation between isogenic strains that lacked CheY and had either low or high levels of fumarate. The difference in the fumarate levels was due to a deletion of the genes encoding the enzymes that synthesize and metabolize fumarate; succinate dehydrogenase and fumarase, respectively. The strains were in a gutted background (i.e. a background deleted for the cytoplasmic chemotaxis proteins and some of the receptors), and switching was achieved by carrying out the measurements at 2.5 degreesC, where it has been demonstrated that gutted cells switch spontaneously. The flagellar rotation of the strain with the highest level of fumarate was the most clockwise-biased and had the highest reversal frequency, indicating that fumarate is effective even in the absence of CheY. Fumarate reduced the free energy difference of the counterclockwise-to-clockwise transition and had no appreciable effect on the activation energy of this transition. Similar observations were made at room temperature, provided that intracellular CheY was present. In a wild-type background, both mutants made rings on semi-solid agar typical of normal chemotaxis. Taken together, the results suggest that the target of fumarate is the switch-motor complex, that fumarate acts by increasing the probability of the clockwise state, and that a fumarate level as low as that found in succinate dehydrogenase mutants is sufficient for normal chemotaxis.

Bacterial Proteins↗
PubMed1998

Reduction of hypercholesterolemic atherosclerosis by CDC-flaxseed with very low alpha-linolenic acid.

Flaxseed (Type I flaxseed) with 51-55% alpha-linolenic acid in its oil and richest source of plant lignans, has been shown to reduce hypercholesterolemic atherosclerosis by 46% without lowering serum lipids. Antiatherogenic activity was claimed to be due to its alpha-linolenic acid and/or lignan content. If alpha-linolenic acid component of flaxseed is responsible for antiatherogenic activity, then, CDC-flaxseed (Type II flaxseed) which has similar oil and lignan content but has very little (2-3% of the total oil) alpha-linolenic acid would have no antiatherogenic effect. An investigation, therefore, was made of Type II flaxseed on high cholesterol diet-induced atherosclerosis and serum lipids [total cholesterol (TC), triglycerides (TG), high-density lipoprotein cholesterol (HDL-C), low-density lipoprotein cholesterol (LDL-C), very low-density lipoprotein cholesterol (VLDL-C)] in rabbits. Rabbits were assigned to four groups: Group I, Control; Group II, Type II flaxseed diet (7.5 g/kg orally daily); Group III, 1% cholesterol diet; Group IV, 1% cholesterol diet supplemented with Type II flaxseed (7.5 g/kg orally daily). Blood samples were collected before (0 time) and after 4 and 8 weeks of experimental diets for measurement of serum lipids. Aorta was removed at the end of 8 weeks for assessment of atherosclerotic plaques. Serum TC, LDL-C, TC/HDL-C, and LDL-C/HDL-C were lower in Group IV as compared to Group III by 14 and 31%, 17 and 32%, 28 and 34% and 24 and 32%, respectively, at 4 and 8 weeks. HDL-C was not affected by Type II flaxseed in hypercholesterolemic rabbit. TG and VLDL-C were markedly increased in Group IV as compared to Group III. Type II flaxseed reduced the development of atherosclerosis by 69%. Histological changes in the atherosclerotic regions were qualitatively similar in Groups III and IV. Results indicate that reduction in hypercholesterolemic atherosclerosis by Type II flaxseed is due to a decrease in serum TC and LDL-C. In conclusion, antiatherogenic activity of Type II flaxseed is not due to alpha-linolenic acid.

Animals↗
PubMed1998

Increased oxidative stress in rat liver and pancreas during progression of streptozotocin-induced diabetes.

1. Oxygen free radicals have been suggested to be a contributory factor in complications of diabetes mellitus. There are many reports indicating the changes in parameters of oxidative stress in diabetes mellitus. In this study we aimed to identify whether oxidative stress occurs in the liver and pancreas in the initial stages of development of diabetes. 2. We therefore investigated the lipid peroxide level (thiobarbituric acid-reactive substances, TBARS) and activities of antioxidant enzymes [superoxide dismutase (SOD), catalase and glutathione peroxidase] in liver and pancreas of control and streptozotocin-induced diabetic rats at various stages of development of diabetes. 3. Male Sprague-Dawley rats were divided into two groups: group I, control (n = 42) and group II, diabetic (n = 42). Each group was further subdivided into seven groups consisting of six rats each. Rats in these subgroups were studied at weekly intervals (0 to 6 weeks). Plasma glucose levels, TBARS levels and activities of antioxidant enzymes were measured in liver and pancreas at various time intervals. 4. There was a significant (P < 0.05) and progressive increase in TBARS levels of liver and pancreas in the diabetic group. Total SOD and Cu-Zn-SOD activity increased (P < 0.05) with progression of diabetes while Mn-SOD activity showed no significant change in either tissue. Catalase and glutathione peroxidase activities increased significantly (P < 0.05) in liver and pancreas. 5. Immunohistochemical study of pancreatic islet revealed a decrease in the expression of insulin with progression of diabetes. However, glucagon and somatostatin showed an increase in immunoreactivity and a difference in their distribution pattern. 6. The findings of the present study suggest that oxidative stress starts at early onset of diabetes mellitus and increases progressively. In conclusion, the structural damage to these tissues or complications of diabetes mellitus may be due to oxidative stress.

Animals↗
PubMed1998

Dipyridamole stress thallium-201 perfusion abnormalities in patients with hypertrophic cardiomyopathy. Relationship to clinical presentation and outcome.

AIMS: Thallium-201 perfusion abnormalities are common in patients with hypertrophic cardiomyopathy and may be associated with an adverse prognosis in the young. The aim of this study was to prospectively determine the relationship between thallium-201 defects during dipyridamole stress to clinical presentation and outcome in a large consecutive series of patients with hypertrophic cardiomyopathy. METHODS/RESULTS: Thallium-201 single photon computed tomography was performed in 216 patients with hypertrophic cardiomyopathy during dipyridamole stress (0.5 mg. kg-1). Fixed perfusion defects occurred in 25%, and reversible defects in 22%. A combination of defects was present in 7%. Fixed defects were associated with: a history of syncope (17 of 46 with, vs 36 of 170 without syncope, P = 0.03); larger left ventricular end-diastolic (46.9 +/- 7.4 mm vs 43.3 +/- 6.4 mm; P = 0.001) and end-systolic dimension (30.2 +/- 8.4 mm vs 24.5 +/- 5.9 mm, P < 0.0001); increased left atrial diameter (46.1 +/- 8.1 mm vs 40.5 +/- 7.7 mm, P < 0.0001); lower fractional shortening (35.9 +/- 10.4% vs 43.8 +/- 8.6%, P < 0.0001): and lower maximal exercise oxygen consumption (24.2 +/- 8.1 ml. min-1. kg-1 vs 29.4 +/- 8.8 ml. min-1. kg-1, P < 0.0003). Reversible defects did not correlate with symptomatic status, but were associated with: larger left atrial dimensions (44.5 +/- 8.1 mm vs 41.0 +/- 8.0 mm; P = 0.009) and greater maximal left ventricular wall thickness (24.0 +/- 7.0 mm vs 20.6 +/- 7.0 mm, P = 0.003). The mean follow up time was 41 +/- 21 months, range 0.6-124. There was no association between any thallium-201 abnormality and disease related death in young or adult patients. CONCLUSION: The present study shows that fixed thallium-201 perfusion defects detected during dipyridamole stress in patients with hypertrophic cardiomyopathy are associated with syncope, larger left ventricular cavity dimensions and reduced exercise capacity. Although the event rate was relatively small, there was no evidence for an association between thallium-201 defects and survival.

Adolescent↗
PubMed1998

Initiating sequences in exercise induced idiopathic ventricular tachycardia of left bundle branch-like morphology.

Initiating sequences for VT may infer the underlying arrhythmogenic mechanisms. This study examines the initiating sequences of exercise induced idiopathic VT of left bundle branch block-like (LBBB-like) morphology and makes an attempt to relate these to clinical aspects and the mechanisms of arrhythmia. Thirty-two patients (mean age 33.4 +/- 13.2 years; 18 men) with exercise induced VT in the absence of structural cardiac abnormality on history, clinical examination, and noninvasive and invasive investigations were divided into two groups on the basis of the initiating sequence of VT on exercise. Group I consisted of patients with long-short sequence of RR intervals prior to the onset of VT (initiating/preinitiating cycle length ratio < or = 0.78). Group II consisted of patients without changes in cycle length prior to VT. Group I mechanism would suggest delayed afterdepolarizations (DADs) or reentry whereas group II mechanism triggered activity due to early afterdepolarizations. Fourteen patients (group I) had long-short sequence and 18 patients (group II) were without cycle length changes prior to VT initiated during exercise. VT axis was inferior in all 18 patients in group II but only in 9 patients in group I (P = 0.02). In these predefined patient groups, sustained monomorphic VT could not be initiated by programmed stimulation in any patient in group I, whereas four patients in group II had inducible VT. Patients in group II also had higher incidence of sustained VT on ambulatory monitoring (P < 0.05). The two groups did not differ in other respects. This study demonstrates the existence of at least two possible mechanisms of initiation of exercise induced idiopathic VT of LBBB-like morphology. VT initiated without cycle length changes is more common, more likely to have an inferior axis suggesting an outflow tract origin, and is probably related to triggered activity secondary to DADs. VT initiated with a long-short sequence is more often nonsustained and may have a superior axis suggesting an origin from the body or septal region of the ventricle. The two groups, therefore, exhibit differences in electrophysiological characteristics that may aid classification and therapy of this arrhythmia.

Adult↗
PubMed1998

T wave complexity in patients with hypertrophic cardiomyopathy.

The complexity of the T wave assessed by principal component analysis (PCA) has been proposed to reflect abnormal repolarization, which may be arrhythmogenic. To determine whether PCA can differentiate patients with hypertrophic cardiomyopathy (HCM) from normal subjects and whether PCA is of prognostic importance in HCM, 112 patients with HCM (41 +/- 14 years, 64 males) and 72 healthy subjects (39 +/- 9 years, 41 males) were studied. Patients with sinus node dysfunction, AV conduction block, flat T waves, QRS > 140 ms, and those < 15 years were excluded from this study. Standard 12-lead ECGs were recorded digitally using the MAC-VU system (Marquette Medical Systems). PCA parameters were computed using the QT Guard software package by Marquette. PCA ratio was significantly greater in HCM patients than in normal controls (23.9% +/- 12.4% vs 16.1% +/- 7.6%, P < 0.0001) and was correlated with QT-end dispersion (r = 0.24, P = 0.01) and QT peak (Q point to T peak) dispersion (r = 0.35, P < 0.0001). HCM patients with syncope (n = 23) had increased PCA ratios compared with those without syncope (29.1% +/- 11.5% vs 22.5% +/- 12.3%, P = 0.01). PCA ratio was similar in patients with and without nonsustained ventricular tachycardia on Holter (25.9% +/- 11.4% vs 22.7% +/- 12.1%, P = 0.2), as well as in patients treated with amiodarone or sotalol versus those not on therapy. In conclusion, assessment of the complexity of the T wave by PCA differentiates HCM patients from normal subjects. PCA ratio correlated with QT dispersion and an increased PCA ratio was associated with a history of syncope in HCM.

Adult↗
PubMed1998

Sudden death in hypertrophic cardiomyopathy: potential importance of altered autonomic control of vasculature.

Current evidence suggests that alterations in the autonomic function and abnormal vascular control play a significant role either as independent triggers themselves or as modifiers of ischaemia and tolerance to to arrhythmias. A combination of several factors--that is, arrhythmia, hypotension, altered autonomic function including vascular control, and ischaemia are therefore likely to act as triggers for sudden death. The relative contribution of each of these factors needs further detailed study.

Adolescent↗
PubMed1998

Oxygen Free Radicals and Endotoxic Shock: Effect of Flaxseed.

BACKGROUND: Cardiac dysfunction and tissue injury during endotoxemia may be caused by increased levels of oxygen free radicals. METHODS AND RESULTS: We therefore investigated the effects of endotoxic shock on cardiac function and contractility, plasma creatine kinase (CK) activity and lactate concentration, oxyradical-producing activity of polymorphonuclear leukocytes (PMNL-CL) and white blood corpuscles, antioxidant reserve (cardiac chemiluminescence [LV-CL]), antioxidant enzyme activity (superoxide dismutase, catalase, glutathione peroxidase), cardiac malondialdehyde (MDA) concentration, a lipid peroxidation product, and hemodynamics in the absence or presence of flaxseed treatment in anesthetized dogs. Flaxseed contains lignans that have antioxidant activites and inhibit platelet-activating factor (PAF). The dogs were assigned to three groups: group I, sham control; group II, endotoxin (ET) treated (5 mg/kg intravenously); group III, ET + flaxseed (2 gm/kg/day orally) for 6 days. ET produced a decrease in cardiac function and contractility and antioxidant enzyme levels, and an increase in cardiac MDA and LV-CL, PMNL-CL, and plasma CK and lactate. Pretreatment with flaxseed attenuated the ET-induced cardiac dysfunction and cellular damage. Protection was incomplete for cardiovascular function, plasma CK, and lactate. CONCLUSIONS: These results suggest that oxyradicals and/or PAF may be involved in the deterioration of cardiovascular function and cellular integrity during ET shock and that antioxidant and anti-PAF agents may be effective in the treatment of ET shock.

Journal Article↗
PubMed1997

Prospective prognostic assessment of blood pressure response during exercise in patients with hypertrophic cardiomyopathy.

BACKGROUND: Previous studies revealed that an abnormal blood pressure response (ABPR) during exercise was common in young hypertrophic cardiomyopathy (HCM) patients and was associated with a family history of premature sudden cardiac death (SCD). This study was performed prospectively to assess the prognostic significance of blood pressure response during exercise in young patients with HCM. METHODS AND RESULTS: Maximum symptom-limited treadmill exercise testing with continuous blood pressure monitoring was performed in 161 consecutive patients 8 to 40 years old (27+/-9). A normal blood pressure response, defined as an increase in the systolic pressure of at least 20 mm Hg from rest to peak exercise in the absence of a fall of >20 mm Hg from peak pressure, was seen in 101 (63%). In 60 (37%), the blood pressure response was abnormal. There was no significant difference in patients with normal blood pressure response and ABPR in terms of age, sex, follow-up, or recognized risk factors for SCD. During the follow-up period (mean, 44+/-20 months), SCD occurred in 12 patients: 3 (3%) in the normal blood pressure response group versus 9 (15%) in the ABPR group (P<.009). ABPR had a sensitivity of 75%, a specificity of 66%, a negative predictive value of 97%, and a positive predictive value of 15% for the prediction of SCD. There was no significant difference in the incidence of other recognized risk factors between patients with SCD and the survivors. CONCLUSIONS: A normal exercise blood pressure response identifies low-risk young patients with HCM. An ABPR identifies the high-risk cohort; the low positive predictive accuracy, however, indicates that further risk stratification is warranted.

Adolescent↗
PubMed1997

Association of angiotensin-converting enzyme gene I/D polymorphism with change in left ventricular mass in response to physical training.

BACKGROUND: The absence (deletion allele [D]) of a 287-base pair marker in the ACE gene is associated with higher ACE levels than its presence (insertion allele [I]). If renin-angiotensin systems regulate left ventricular (LV) growth, then individuals of DD genotype might show a greater hypertrophic response than those of II genotype. We tested this hypothesis by studying exercise-induced LV hypertrophy. METHODS AND RESULTS: Echocardiographically determined LV dimensions and mass (n=140), electrocardiographically determined LV mass and frequency of LV hypertrophy (LVH) (n=121), and plasma brain natriuretic peptide (BNP) levels (n=49) were compared at the start and end of a 10-week physical training period in male Caucasian military recruits. Septal and posterior wall thicknesses increased with training, and LV mass increased by 18% (all P<.0001). Response magnitude was strongly associated with ACE genotype: mean LV mass altered by +2.0, +38.5, and +42.3 g in II, ID and DD, respectively (P<.0001). The prevalence of electrocardiographically defined LVH rose significantly only among those of DD genotype (from 6 of 24 before training to 11 of 24 after training, P<.01). Plasma brain natriuretic peptide levels rose by 56.0 and 11.5 pg/mL for DD and II, respectively (P<.001). CONCLUSIONS: Exercise-induced LV growth in young males is strongly associated with the ACE I/D polymorphism.

Adult↗
PubMed1997

Dietary flax seed in prevention of hypercholesterolemic atherosclerosis.

Oxygen free radicals (OFRs) have been implicated in the development of hypercholesterolemic atherosclerosis. Flax seed is the richest source of omega-3 fatty acid and lignans. omega-3 Fatty acid suppresses the production of interleukin-1 (IL-1), tumor necrosis factor (TNF) and leukotriene B4 (LTB4), and of OFRs by polymorphonuclear leukocytes (PMNLs) and monocytes. Lignans possess anti-platelet activating factor (PAF) activity and are antioxidant. PAF, IL-1, TNF and LTB4 are known to stimulate PMNLs to produce OFRs. Flaxseed would, therefore, reduce the levels of OFRs and hence would prevent the development of hypercholesterolemic atherosclerosis. The effects of dietary flax seed on a high cholesterol diet induced atherosclerosis, lipid profile and OFR-producing activity of PMNLs (PMNL-CL) were investigated in rabbits. The rabbits were divided into 4 groups: group I, control; group II, flax seed diet (7.5 g/kg daily, orally); group III, 1% cholesterol diet; and group IV, same as group III but received flax seed (7.5 g/kg daily, orally). Blood samples were collected before and after 4 and 8 weeks on their respective diets for biochemical measurements and aortae were removed at the end of 8 weeks for estimation of atherosclerotic changes. The high cholesterol diet increased the serum level of total cholesterol (TC) and PMNL-CL without altering the levels of serum triglycerides (TG). These changes were associated with a marked development of atherosclerosis in the aorta. Flax seed reduced the development of aortic atherosclerosis by 46% and reduced the PMNL-CL without significantly lowering the serum cholesterol. Flax seed in normocholesterolemic rabbits increased serum total cholesterol and decreased PMNL-CL without significantly affecting the serum TG. Modest dietary flax seed supplementation is effective in reducing hypercholesterolemic atherosclerosis markedly without lowering serum cholesterol. Its effectiveness against hypercholesterolemic atherosclerosis could be due to suppression of enhanced production of OFRs by PMNLs in hypercholesterolemia. Dietary flax seed supplementation could, therefore, prevent hypercholesterolemia-related heart attack and strokes.

Animals↗
PubMed1997

Auxilin-induced interaction of the molecular chaperone Hsc70 with clathrin baskets.

We previously reported that a 100-kDa cofactor, recently identified as auxilin, is a DnaJ homolog which is required for Hsc70 to uncoat clathrin baskets. In the present study we investigated the effect of auxilin on the interaction of Hsc70 with pure clathrin baskets at pH 6, where no uncoating occurs. In a reaction which required auxilin, the baskets activated the Hsc70 ATPase activity more than 100-fold with an apparent dissociation constant of about 0.2 microM. Maximal ATPase activity occurred at a 1 to 1 molar ratio of auxilin to clathrin triskelion independent of the Hsc70 concentration suggesting that auxilin is primarily complexed with the clathrin baskets. The binding of Hsc70 to baskets also required auxilin, but less auxilin was needed for maximum binding than for maximum ATPase activity showing that auxilin can catalytically induce binding of Hsc70. The binding also required ATP; Hsc70 dissociated from baskets with a 6 min half-life when ATP was hydrolyzed to ADP. In contrast to auxilin, the assembly proteins, AP-2 and AP180, did not support activation of the Hsc70 ATPase activity by clathrin baskets nor did soluble clathrin triskelions at pH 7 significantly activate the ATPase activity with auxilin present. Therefore, the interaction of auxilin, clathrin baskets, and Hsc70-ATP is highly specific with auxilin first binding to a clathrin triskelion in the baskets and then Hsc70-ATP strongly binding to the auxilin-clathrin complex; the auxilin can then migrate to another clathrin triskelion before the ATPase cycle is complete.

Adaptor Proteins, Vesicular Transport↗